eEF1A1 promotes colorectal cancer progression and predicts poor prognosis of patients

Abstract

Colorectal cancer (CRC) is a major leading cause of cancer mortality worldwide in which dysregulated protein synthesis plays an etiologic role. The eukaryotic elongation factor 1 A1 (eEF1A1) exerts significant effects on protein synthesis by contributing to peptide chain extension. Whereas its role in CRC remains to be investigated. In this study, we found that the mRNA and protein levels of eEF1A1 were significantly upregulated in CRC cell lines and tissues. Elevated expression of eEF1A1 was correlated with shorter overall survival in 94 CRC patients. The inhibition of proliferation and cell cycle block were observed in CRC cells after eEF1A1 downregulation. Mechanistically, weighted gene correlation network analysis and further Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis suggested that mitogen-activated protein kinases (MAPKs) signaling pathways were significantly enriched in high-eEF1A1 expression group, and the levels of phosphorylated p38/JNK/ERK MAPK were dramatically decreased after eEF1A1 downregulation. Overexpression of eEF1A1 in CRC correlated with a poor prognosis. Collectively, this study determined the oncogenic role of eEF1A1 in CRC proliferation and tumorigenesis. eEF1A1 might be a promising therapeutic target and prognostic biomarker in CRC.

Keywords: colorectal cancer; eukaryotic elongation factor 1 A1 (eEF1A1); mitogen-activated protein kinase (MAPK); proliferation.

FIGURE 1

Increased eEF1A1 expression in colorectal cancer (CRC) cell lines and tissues. (A) eEF1A1 expression in paired CRC samples from GSE18105 and TCGA database. (B) eEF1A1 expression in CRC samples from GES21510 and GSE44076. (C) The protein levels of eEF1A1 in 14 paired CRC and adjacent non‐tumorous tissues. The fold change accordingly was used to calculate the average eEF1A1 expression levels in CRC and adjacent non‐tumors. The ratio of eEF1A1/GAPDH of the gray values was used to normalize the protein expression of eEF1A1. Quantitative data represent mean ± SD. (D) The mRNA levels of eEF1A1 in 14 pairs of CRC and corresponding adjacent non‐tumor tissues. (E) The mRNA expression levels of eEF1A1 in the NCM460 and CRC cell lines SW480, SW620, LOVO, RKO, Caco2, and HCT8. The assay repeated three times. (F) The protein expression levels of eEF1A1 in NCM460 and CRC cell lines SW480, SW620, LOVO, RKO, Caco2, and HCT8.

FIGURE 2

eEF1A1 promotes colorectal cancer cell proliferation in vitro. (A) siRNAs against eEF1A1 (sieEF1A1) were used to knock down eEF1A1 in RKO and Caco2 cells. The silencing of eEF1A1 were confirmed by Western blot and qRT‐PCR. (B) The cell viability of eEF1A1‐knockdown RKO and Caco2 cells was determined by colony formation assays. (C) The proliferation capacity of eEF1A1‐knockdown RKO and Caco2 cells was detected by CCK‐8 assay. (D) The cell cycle distribution of successfully transfected RKO and Caco2 cells. (E) The protein levels of Cyclin D1, Cyclin E1, CDK2, and CDK4 in eEF1A1‐knockdown RKO and Caco2 cells. (F) The protein levels of Bax, Bcl‐2, Caspase‐3, and Cleaved Caspase‐3 in eEF1A1‐knockdown RKO and Caco2 cells.

FIGURE 3

eEF1A1 induces colorectal cancer tumorigenesis in vivo. (A) Photos of xenograft tumors derived by injecting RKO‐sheEF1A1‐1 cells, RKO‐sheEF1A1‐2 or RKO‐sheEF1A1‐NC cells. (B) The xenograft tumors volume was measured every 3 days, tumor growth curves (left) and tumor weight (right) are shown. (C) HE, IHC staining of eEF1A1 and Ki‐67 in the xenograft are shown. Scale bars, 50 μm.

FIGURE 4

eEF1A1 actives MAPK signaling in colorectal cancer cells. (A) Heatmap of module‐trait relationships. The tan module block is selected for subsequent analysis. (B) KEGG analysis of genes in tan module block. Color depth indicates the enrichment significance, circle size presents the enriched gene count. (C) The protein levels of MAPK signaling pathways related was detected by western blot in eEF1A1‐knockdown RKO and Caco2 cells.

FIGURE 5

Overexpression of eEF1A1 in colorectal cancer (CRC) correlated with a poor outcome. (A) eEF1A1 expression in paired CRC samples from TMA cohort performed. Scale bars, 600 μm (above) and 40 μm (below). Representative images (left) and IHC score grades of eEF1A1 expression are shown (right). (B) Correlation between eEF1A1 expression and OS in a TMA cohort. (C) Correlation between eEF1A1 expression and OS in GSE39582 and GSE17536.